Gurskaya Nadya G, Verkhusha Vladislav V, Shcheglov Alexander S, Staroverov Dmitry B, Chepurnykh Tatyana V, Fradkov Arkady F, Lukyanov Sergey, Lukyanov Konstantin A
Institute of Bioorganic Chemistry, Russian Academy of Sciences, Miklukho-Maklaya 16/10, Moscow 117997, Russia.
Nat Biotechnol. 2006 Apr;24(4):461-5. doi: 10.1038/nbt1191. Epub 2006 Mar 19.
Green fluorescent protein (GFP) and GFP-like proteins represent invaluable genetically encoded fluorescent probes. In the last few years a new class of photoactivatable fluorescent proteins (PAFPs) capable of pronounced light-induced spectral changes have been developed. Except for tetrameric KFP1 (ref. 4), all known PAFPs, including PA-GFP, Kaede, EosFP, PS-CFP, Dronpa, PA-mRFP1 and KikGR require light in the UV-violet spectral region for activation through one-photon excitation--such light can be phototoxic to some biological systems. Here, we report a monomeric PAFP, Dendra, derived from octocoral Dendronephthya sp. and capable of 1,000- to 4,500-fold photoconversion from green to red fluorescent states in response to either visible blue or UV-violet light. Dendra represents the first PAFP, which is simultaneously monomeric, efficiently matures at 37 degrees C, demonstrates high photostability of the activated state, and can be photoactivated by a common, marginally phototoxic, 488-nm laser line. We demonstrate the suitability of Dendra for protein labeling and tracking to quantitatively study dynamics of fibrillarin and vimentin in mammalian cells.
绿色荧光蛋白（GFP）及类GFP蛋白是非常重要的基因编码荧光探针。在过去几年里，一类新型的可光激活荧光蛋白（PAFP）被开发出来，这类蛋白能产生显著的光诱导光谱变化。除了四聚体KFP1（参考文献4），所有已知的PAFP，包括PA-GFP、Kaede、EosFP、PS-CFP、Dronpa、PA-mRFP1和KikGR，都需要紫外-紫光光谱区域的光通过单光子激发来激活——这种光对某些生物系统可能具有光毒性。在此，我们报道了一种单体PAFP，Dendra，它源自八放珊瑚树状软珊瑚（Dendronephthya sp.），在可见光蓝光或紫外-紫光照射下，能够实现1000至4500倍的从绿色荧光态到红色荧光态光转换。Dendra是首个同时具备单体性质、在37摄氏度下能有效成熟、激活态具有高光稳定性且能用常见的、光毒性较小的488纳米激光线进行光激活的PAFP。我们证明了Dendra适用于蛋白质标记和追踪，以定量研究哺乳动物细胞中纤维原蛋白和波形蛋白的动态变化。
